Project Summary At the molecular level atherosclerosis can be defined as an assembly of hundreds of intra- and extra-cellular proteins that jointly alter cellular processes and remodel the local environment in characteristic ways. Proteomic changes produce vascular lesions responsible for ischemic cardiovascular events worldwide. Unfortunately, current methods to treat and prevent cardiovascular disease focus on risk factors that are not deterministic of these proteomic changes; or on anatomic manifestations of the disease that are evident long after the proteomic changes are underway. To detect disease early and interrupt the disease process before clinical consequences occur, it is necessary to recognize and understand the specific patterns of arterial proteins that constitute the molecular signature of atherosclerosis. Indeed, if these arterial protein changes could be reliably detected and reversed, the possibility of eradication of coronary, cerebral and peripheral arterial disease would be within our grasp. Thus, the long-term goal of our research group is to comprehensively define the proteomic architecture of atherosclerosis, to develop reliable methods to detect these changes, and ultimately to identify mechanisms to prevent or reverse them. In the initial funding (GPAA Phase 1, 2012-2016) our group capitalized on advances in applied proteomics and new analytic methods to develop a clinical, technical and analytic platform to study the human arterial proteome. This work produced clear evidence that early atherosclerotic plaques are characterized by complex changes in the proteomic topology of several cell regulatory pathways. Our current goal is to extend this work to include robust descriptions of the proteomic architecture of atherosclerosis specifically in women and African-Americans, to expand the portfolio of protein features, and develop a multi-dimensional clinical assay that could be used for early detection. To achieve this goal we propose the following Specific Aims: 1. Validate and expand the atherosclerosis-associated protein profiles identified in GPAA in additional autopsy specimens and determine if these profiles vary as a function of sex or ethnicity. 2. Identify protein isoforms and selected PTM proteins that are enriched in atherosclerotic plaques and define significant downstream protein networks. 3. Based on our data, develop and produce a plasma protein biomarker panel to predict the presence of subclinical atherosclerosis and risk for future cardiovascular events. The resulting data will provide novel insights into the pathogenesis of atherosclerosis and support the development of a new clinical tool that should be useful for improved prediction and pre-clinical assessment of therapeutic efficacy of current and novel future interventions.